Abstract
BackgroundFourier transform infrared spectroscopy (FTIR) has been used as analytical tool in chemistry for many years. In addition, FTIR can also be applied as a rapid and non-invasive method to detect and identify microorganisms. The specific and fingerprint-like spectra allow - under optimal conditions - discrimination down to the species level. The aim of this study was to develop a fast and reproducible non-molecular method to differentiate pure samples of Bacillus spores originating from different species as well as to identify spores in a simple matrix, such as the clay mineral, bentonite.ResultsWe investigated spores from pure cultures of seven different Bacillus species by FTIR in reflection or transmission mode followed by chemometrical data treatment. All species investigated (B. atrophaeus, B. brevis, B. circulans, B. lentus, B. megaterium, B. subtilis, B. thuringiensis) are typical aerobic soil-borne spore formers. Additionally, a solid matrix (bentonite) and mixtures of benonite with spores of B. megaterium at various wt/wt ratios were included in the study. Both hierarchical cluster analysis and principal component analysis of the spectra along with multidimensional scaling allowed the discrimination of different species and spore-matrix-mixtures.ConclusionsOur results show that FTIR spectroscopy is a fast method for species-level discrimination of Bacillus spores. Spores were still detectable in the presence of the clay mineral bentonite. Even a tenfold excess of bentonite (corresponding to 2.1 × 1010 colony forming units per gram of mineral matrix) still resulted in an unambiguous identification of B. megaterium spores.
Highlights
Fourier transform infrared spectroscopy (FTIR) has been used as analytical tool in chemistry for many years
Baldauf and coworkers found attenuated total reflection (ATR) to give better resolution of peaks while using less biomass compared to other FTIR methods [18]
Bacillus species used in this study are typical representatives of aerobic soil-borne spore formers and have been isolated from various soil environments
Summary
Fourier transform infrared spectroscopy (FTIR) has been used as analytical tool in chemistry for many years. FTIR can be applied as a rapid and non-invasive method to detect and identify microorganisms. The aim of this study was to develop a fast and reproducible non-molecular method to differentiate pure samples of Bacillus spores originating from different species as well as to identify spores in a simple matrix, such as the clay mineral, bentonite. Because each different material is a unique combination of atoms, no two compounds produce exactly the same infrared. In 1911, W.W. Coblentz (cited by [1]) was the first to analyze complex biological samples such as gelatine or chitin as well as minerals (e.g., quartz, opal, and muscovite) by infrared spectroscopy. Naumann and coworkers [9] developed efficient methods and instruments for microbiological use in a joint project between the Robert Koch Institute and a manufacturer of FTIR spectrometers (Bruker, Germany)
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